Animal cells have receptors on the outer surface which bind molecules having specific receptor binding domains, resulting in an effect on the biological function of the cells. If this function is incompatible with the well being of the host, these receptor specific molecules could be employed to deliver therapeutic agents to the cell if the agents could be attached to such molecules without destroying the ability of the molecule to bind to the cell surface.
Abnormal, and in some situations even normal functioning of particular cell types may precipitate life threatening or other disease conditions. In immune system dysfunction, the inappropriate activity of T lymphocytes (T cells) plays a particularly prominent part, and therapy now frequently consists of treatment with strong immunosuppressive agents. However, many of these treatments result in widespread toxicity and resulting damage to the immune system frequently extends beyond the particular cell type causing the disorder. Thus, the recovering patient may be left temporarily stripped of defenses against infectious pathogens. A better therapy for conditions such as autoimmune disease and graft rejection would be one that concentrates only on those clones of T cells that are activated by inappropriate targets, leaving intact the remaining T lymphocytes and other cells of the immune system.
An important feature that distinguishes activated from resting T cells is the expression of high affinity receptors for interleukin-2 (IL-2). There are two IL-2 receptors, high- and low affinity. The low affinity receptor is found on both resting and activated T cells, while the high affinity receptor is found only on activated cells and is the receptor involved in the internalization of interleukin-2.
After contact with cellular antigens that they recognize, certain T cells secrete lymphokines, including interleukin-2, that mediate a variety of immune functions. In addition, all T cells respond to antigen activation by expressing receptors for interleukin-2, thereby becoming responsive to the powerful growth-promoting effects of this hormone. The resulting proliferation amplifies the number of cells available to react to the current antigenic challenge. When the antigen is cleared, the number of high affinity interleukin-2 receptors is reduced.
Targeting of particular cell classes, including cells of the immune system, has been attempted. In particular, monoclonal antibodies (MoAbs) against the human low affinity interleukin-2 receptor are available, and some groups have explored the use of such monoclonal antibodies coupled to cytotoxic agents to target interleukin-2 receptor-bearing leukemic T cells for destruction.
Liposomes with antibodies attached to their surfaces have been shown to bind specifically to cells expressing a cell surface antigen. A problem in delivering cytotoxic agents as conjugates with antibodies against the interleukin-2 receptor is that the antibody is not internalized. The high affinity interleukin-2 site is primarily associated with the internalization of interleukin-2. As one result of the low frequency of internalization of antibodies, impractically high concentrations of antibody-toxin conjugate must be used, with a corresponding increased risk of toxicity to bystander (non-interleukin-2 receptor bearing) cells.
While it is known to covalently link certain antibodies to liposomes subsequent to the random partial protection of --NH.sub.2 functional groups of the antibody (Jansons, et al., 1981 Anal. Biochem. 111, 54), attempts to couple receptor specific proteins to therapeutic agents or liposomes in order to achieve targeted delivery to particular cell populations have not been successful due to the fact that the existing coupling processes have failed to provide a means to ensure that the receptor binding domain on such a protein is not destroyed, and thus remains accessible for later interaction with cell receptor sites.
Although chemical techniques are available for linking proteins to liposomes, these techniques are nonselective in nature and involve linkages which will block or otherwise interfere with the receptor binding domain on the protein. Thus, it has been a desideratum to provide a liposome having a protein hormone with active receptor binding domains attached thereto, and means for binding protein hormones such as interleukin-2 to hposomes while protecting the receptor binding domain of the protein.